Influenza infection remains an enormous public health concern despite the availability of vaccines and worldwide surveillance. The high mutation rates of influenza A viruses (IAVs) enable the viruses to evade natural and vaccine-mediated immunity. Also, current anti-viral therapies do not prevent IAV-related deaths when there is a delay in initiating treatment. Thus, new therapeutic options for treating influenza disease are an immediate public health priority. Our pilot studies identify MMP-9 as an attractive IAV therapeutic target. MMP- 9 is strikingly upregulated in plasma samples from human subjects infected with seasonal and H1N1 IAV and in lungs from mice infected with H1N1 IAV. In mice, Mmp-9 increases IAV-associated mortality and late-stage lung inflammation and late-stage lung viral burdens. We proposed to test the therapeutic efficacy of a re- purposed therapeutic candidate (ADZ1236) that selectively and potently inhibits MMP-9 activity in IAV infections in two species in three integrated and highly collaborative aims. Aim 1 will test the therapeutic efficacy of AZD1236 therapy in mice infected with BL2 H1N1 IAV. AZD1236 will be tested in mice alone and in combination with a clinically-used antiviral agent (the neuraminidase inhibitor, oseltamivir). A delayed initiation of treatment approach will be optimized to model rea-world treatment scenarios for serious IAV infections. Aim 2 will identify the mechanisms by which Mmp-9 promotes (and AZD1236 limits) serious IAV infections in mice. We will study H1N1-infected WT vs. Mmp-9-/- mice and Mmp-9 bone marrow chimeric mice and use in vitro approaches to test our hypothesis that Mmp-9 promotes adverse outcomes in IAV disease by cleaving host or viral proteins. These Mmp-9 substrates will be identified. Aim 3 will test the therapeutic efficacy of AZD1236 in ferrets infected with BL2 H1N1 and the BL3 highly pathogenic avian influenza (HPAI) H5N1 strain (and in H7N9 IAV-infected ferrets if this strain emerges as an epidemic or pandemic strain during the funding period). Our studies will determine whether AZD1236-mediated MMP-9 inhibition has therapeutic efficacy against IAV in both small and large animals sufficient to thereby satisfy the FDA Two Animal Rule required to demonstrate efficacy to apply for FDA approval. Successful completion of the work proposed herein may provide a first in class therapeutic intervention for serious influenza-mediated lung disease.